Passalora fulva (Cooke) U. Braun & Crous [syn: Cladosporium fulvum Cooke, Fulvia fulva (Cooke) Ciferri] is a biotrophic pathogen that causes leaf mold of tomato (Solanum lycopersicum L.) (Braun et al. 2003; Crous and Braun 2003). The disease is a primary problem on greenhouse tomatoes and occurs around the world when relative humidity is high. Genes conferring resistance to P. fulva (Cf-2, Cf-4, Cf-5, Cf-6, Cf-9, and Cf-11) have been introduced into cultivated tomato from wild species, and these resistance genes show a typical gene-for-gene relationship (Thomma et al. 2005). However, new races that overcome the resistance genes have rapidly evolved, and isolates that can overcome all resistance genes except Cf-6 have appeared in Europe since the 1980s (Lindhout et al. 1989).

In Japan, the resistance of cultivars that carry Cf-2, Cf-4, Cf-9, and Cf-11 has been overcome by eight races of P. fulva (Enya et al. 2009; Kishi 1962; Ozaki and Shirakawa 1996; Satou et al. 2005; Yamada and Abiko 2002). Races that have overcome Cf-9 resistance first occurred in Gunma, Chiba, and Fukushima prefectures in 2007 and were identified as races 4.9 and 4.9.11 (Enya et al. 2009). In 2008, leaf mold was also found on Cf-9 tomato cultivars in Iwate prefecture, geographically distant from the area where races 4.9 and 4.9.11 have occurred. The present study was undertaken to determine whether the 2008 outbreak involved the same races or a new race had developed.

Disease symptoms are usually limited to the foliage and first appear on the upper surfaces of leaves as pale green or yellow spots with indefinite margins (Fig. 1a). The lower surfaces of the patches become covered with a velvety, olive-brown fungus growth (Fig. 1b). As the disease progresses, the leaf spots turn yellowish brown. The leaves curl, wither, and drop prematurely. Defoliation starts at the bottom of the plant and progresses upward. In 2008, five single-spore isolates were obtained from cvs. ‘Momotaro-Sunny’ and ‘Momotaro-Natsumi’ (Takii & Co., Kyoto, Japan), hybrid cultivars that carry Cf-9. Their conidia were catenate, acropleurogenous, pale to dark brown, cylindrical or ellipsoid, straight or mildly curved, had 0 to 2 septa (mostly 0 or 1), and were 9–29 × 4–8 μm in size on potato dextrose agar (Difco, Detroit, MI, USA; Fig. 1c). Disease symptoms and the morphological characteristics of these isolates were similar to those described previously for P. fulva (Holiday and Mulder 1976).

Fig. 1
figure 1

Symptoms and causal organism of leaf mold of tomato. a Typical symptom of a tomato leaf was yellowish spots with indefinite margins on the upper surface of tomato leaf. b On the corresponding areas of the lower leaf surface, the fungus appears as an olive green velvety growth with abundant conidia. c Conidia produced on potato dextrose agar (PDA). Bar 20 μm

Full size image

To determine the race of the isolates, we tested them on a set of differential genotypes that contained either one or two resistance genes. The tomato cultivars ‘Potentate’ (no resistance gene), ‘Vetomold’ (Cf-2), ‘Purdue 135’ (Cf-4), ‘Moneymaker-Cf-5’ (Cf-5), ‘Ontario 7818’ (Cf-6), ‘Moneymaker-Cf-9’ (Cf-9), and ‘Ontario 7716’ (Cf-4 and Cf-11) were used for all inoculations. Isolates were cultured in the dark on potato dextrose agar (Difco) for 2 weeks at 25°C. Three 4-week-old plants of each cultivar were sprayed on the lower side of the leaves with a conidial suspension (104 conidia/mL). The inoculated plants were incubated in a moist chamber at 100% relatively humidity and 25°C with a 12 h light/12 h dark photoperiod. After 2 weeks, plants were checked for symptoms and conidia formation. The inoculated plants were scored visually as either resistant or susceptible based on disease development. Inoculation tests were repeated twice for each strain.

All five isolates were virulent on ‘Potentate’, ‘Vetomold’, and ‘Moneymaker-Cf-9’ and generated a large number of conidia on the upper and lower sides of the leaves (Table 1). Other cultivars showed resistance to all isolates by means of a hypersensitive response. On the basis of the differential interaction of the tomato genotypes with the known Cf genes, we propose designating these isolates as a new race 2.9 that has not been reported previously. Recent studies showed that new races of P. fulva have appeared within several years of the release and prevalence of new tomato varieties based on Cf resistance genes in Japan (Enya et al. 2009; Satou et al. 2005; Yamada and Abiko 2002). In Iwate prefecture, the Cf-9 varieties have been cultivated since 2006, and new race 2.9 was appeared. Our results suggest that new races of P. fulva develop rapidly and have overcome the resistance provided by new Cf resistance genes within the last several years.

Table 1 Virulence spectrum of races of Passalora fulva isolated in Japan on tomato genotypes
Full size table

Of the nine races of P. fulva in Japan, six have already appeared in Europe (Lindhout et al. 1989), but races 4.9, 4.9.11, and 2.9 are unique to Japan. Race 2.9 appeared in 2008 on Cf-9 tomato grown in Iwate prefecture, from which race 2 was isolated in 1996 (Ozaki and Shirakawa 1996). It is likely that some isolates of race 2 have broken down the Cf-9 resistance; these isolates were designated as race 2.9. We are now focusing on four AVR genes (AVR2, AVR4, AVR4E, and AVR9). The encoded proteins induce a host-specific resistance in tomato that carries the Cf-2, Cf-4, Cf-4E, and Cf-9 genes, respectively (Thomma et al. 2005). Several mutations in the AVR genes that cause a transition from avirulence to virulence (Stergiopoulos et al. 2007). Future experiments will be required to reveal allelic variation in the AVR genes of these indigenous races.